Hybrid E. Coli Confirmed in European Outbreak

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Note that the epidemic in Germany was caused by a highly virulent Shiga-toxin producing strain of E. coli that also contained antibiotic-resistance elements and was distributed on bean sprouts.

Both studies reported on here indicate that applications of new technologies could be of great help to public health authorities in the near future.

Full DNA sequence analysis of the unusually deadly E. coli strain believed responsible for the recent outbreak in Europe has confirmed that it contained a novel hybrid of genetic elements which likely accounted for its virulence.

Researchers from the U.S. and Denmark determined that the outbreak strain had genes for the Shiga toxin 2 and for a variety of other virulence and antibiotic-resistance elements, the combination of which has rarely been seen in a single E. coli organism, according to their report online in the New England Journal of Medicine.

Eric Schadt, PhD, of Pacific Biosciences in Menlo Park, Calif., and colleagues also found that exposing the outbreak strain in vitro to ciprofloxacin caused it to step up expression of the Shiga toxin.

This latter finding "suggests that caution is warranted in the use of certain classes of antibiotics to counteract this newly emerged pathogen," they wrote.

Similar results were also reported online in NEJM by a group based in China. Those researchers sequenced an isolate from the recent outbreak and uploaded the data to an open-source website where it was analyzed against published sequences for other E. coli strains by an informal, volunteer group of bioinformatics specialists working independently.

The outbreak sickened at least 4,000 people and killed 52, mostly in Germany but with a small pocket in western France. Some American travelers to Germany also fell ill after they returned home.

Disease control investigators traced the E. coli contamination to fenugreek seeds imported from Egypt that were used to grow fresh sprouts.

With the last case of infection seen in early July, authorities in Germany this week declared the outbreak officially over.

What made the illnesses unusual was a relatively high rate of hemolytic uremic syndrome as well as the death toll.

In June, two research groups in Germany had found hints that the outbreak strain had virulence factors associated with different classes of E. coli -- a propensity to form colonies on the intestinal wall, production of Shiga toxin, and antibiotic resistance elements.

These suspicions have now been confirmed by the sequence analysis reported in the NEJM.

Schadt and colleagues sequenced the full genome of the O104:H4 C227-11 strain, isolated from a German victim of the outbreak, along with genomes from six other O104:H4 strains and five enteroaggregative strains. They also obtained previous sequence data on three additional O104:H4 isolates.

The researchers also conducted lab experiments with the organisms to connect the genomic data with functional information.

Their findings indicated that the European outbreak strain "is a dramatic example of gene acquisition by means of lateral transfer that resulted in an accretion of synergistic virulence factors," Schadt and colleagues wrote.

"By all molecular definitions, it is an enteroaggregative E. coli strain, but one that has acquired a Shiga-toxin-encoding phage."

The researchers indicated that their data did not conclusively show why hemolytic uremic syndrome was so common with this strain. But they speculated that it stemmed from the organism's large number of serine proteases that facilitate mucosal colonization and damage, in combination with other virulence factors.

Such a mechanism could account for "the increased uptake of Shiga toxin into the circulation," Schadt and colleagues wrote.

The report from the Chinese group, led by Ruifu Yang, MD, PhD, of the Beijing Institute of Microbiology and Epidemiology, suggested that the outbreak strain had originated with an enteroaggregative progenitor that had picked up additional virulence genes.

"Although this outbreak strain has surprised the general public and public health officials, related potential progenitor strains have been reported from three continents," Yang and colleagues wrote.

For example, a 2005 outbreak in Korea with high rates of hemolytic uremic syndrome was traced to an O104:H4 strain, they noted.

Yang and colleagues also identified similarities between the recent European outbreak strain and two others -- one isolated in Germany in 2001 and another from central Africa.

Both research groups highlighted the speed with which they obtained their results, just weeks after the outbreak attracted public attention.

These applications of new technologies -- high-throughput DNA sequencing on the one hand and "crowd-sourced" genomic analysis on the other -- could be of great help to public health authorities in the not-too-distant future, they suggested.

The study by Schadt and colleagues was funded by the National Institutes of Health and the Danish Council for Strategic Research.

The study by Yang and colleagues was supported by the State Key Development Program for Basic Research of China, the National Key Program for Infectious Diseases of China, Shenzhen Biological Industry Development Special Foundation–Basic Research Key Projects, Key Laboratory Project Supported by Shenzhen City, the European Union Microme Program, the Alexander von Humboldt Foundation, the Medical Faculty of the University Medical Center Hamburg–Eppendorf, and the British Biotechnology and Biological Sciences Research Council.

Schadt and many co-authors were employees of Pacific Biosciences. No other relevant financial interests were reported.

One co-author of the report by Yang and colleagues disclosed a travel reimbursement from Pfizer. Other authors including Yang indicated they had no relevant financial interests.

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